DESCRIPTION: The long term goal of this proposal is to understand the biology, biochemistry and genetics of cellular responses to DNA alkylation damage. Alkylating agents represent the most abundant class of chemical DNA damaging agents in our environment and they are even present as natural cellular metabolites. These agents are capable of inducing cell death, mutation, chromosome damage, and cancer. Since we are continuously exposed to these chemicals, and since certain alkylating agents are used for the chemotherapeutic treatment of cancer patients, it is important to understand exactly how cells respond to these agents. The mechanisms that cells employ to defend against DNA damaging agents have been highly conserved, and we now know that bacteria, yeast and mammalian cells employ similar strategies to prevent the induction of cell death and mutation. In recent years Dr. Samson has identified a number of eukaryotic DNA alkylation repair genes encoding DNA glycosylases and DNA methyltransferases. This proposal presents an integrated plan, designed to enhance our understanding of how cells respond to alkylating agents. The plan was designed to encompass research at various levels of development; i.e., at the level of identifying new pathways that protect cells against alkylating agents, at the level of further characterizing recently discovered pathways, and at the level of detailed characterization of pathways that have been under study for several years. This purposefully broad-based plan results in a large, diverse proposal which should enhance our understanding of the eukaryotic response to DNA damage. The Specific Aims include the following: a detailed genetic and biochemical analysis of the transcriptional regulation of the S. cerevisiae MAG 3MeA DNA glycosylase gene and several other DNA repair and metabolism genes; further analysis of the in vivo role MAG; the generation and characterization of cell lines and whole animals (mice) with altered levels of DNA alkylation repair enzymes; the isolation of new eukaryotic genes involved in the response of cells to DNA alkylation damage. The health relatedness of this project lies in the fact that it will contribute to an understanding of some of the events that lead to carcinogenesis and to the appearance of mutations in germ line cells.